The nature and assembly of basement membrane constituents namely IV collagen, laminin and heparan sulphate proteoglycan were studied using a variety of in vitro binding assays. These basement membrane macromolecules were isolated from the EHS tumor grown in C57 black mice. Protease-derived fragments of laminin and IV collagen were characterized by rotary shadowing electron microscopy. The domains required for binding of laminin and IV collagen were identified. Laminin is a cross-shaped molecule with three equal short arms and one long arm. The cell binding region of laminin was also identified and found to reside at the intersection of the three short arms. The carbohydrate composition of laminin was obtained and the distribution of sugars on the long and short arms of laminin molecule was studied.